Electrostatic Contributions of Aromatic Residues in the Local Anesthetic Receptor of Voltage-Gated Sodium Channels

Antiarrhythmics, anticonvulsants, and local anesthetics target voltage-gated sodium channels, decreasing excitability of nerve and muscle cells. Channel inhibition by members of this family of cationic, hydrophobic drugs relies on the presence of highly conserved aromatic residues in the pore-lining S6 segment of the fourth homologous domain of the channel. We tested whether channel inhibition was facilitated by an electrostatic attraction between lidocaine and {pi} electrons of the aromatic rings of these residues, namely a cation-{pi} interaction. To this end, we used the in vivo nonsense suppression method to incorporate a series of unnatural phenylalanine derivatives designed to systematically reduce the negative electrostatic potential on the face of the aromatic ring. In contrast to standard point mutations at the same sites, these subtly altered amino acids preserve the wild-type voltage dependence of channel activation and inactivation. Although these phenylalanine derivatives have no effect on low-affinity tonic inhibition by lidocaine or its permanently charged derivative QX-314 at any of the substituted sites, high-affinity use-dependent inhibition displays substantial cation-{pi} energetics for 1 residue only: Phe1579 in rNaV1.4. Replacement of the aromatic ring of Phe1579 by cyclohexane, for example, strongly reduces use-dependent inhibition and speeds recovery of lidocaine-engaged channels. Channel block by the neutral local anesthetic benzocaine is unaffected by the distribution of {pi} electrons at Phe1579, indicating that our aromatic manipulations expose electrostatic contributions to channel inhibition. These results fine tune our understanding of local anesthetic inhibition of voltage-gated sodium channels and will help the design of safer and more salutary therapeutic agents

A cross sectional study of the prevalence, risk factors and population attributable fractions for limb and body lesions in lactating sows on commercial farms in England

Background: Lesions on sows' limbs and bodies are an abnormality that might impact on their welfare. The prevalence of and risks for limb and body lesions on lactating sows on commercial English pig farms were investigated using direct observation of the sows and their housing. Results: The prevalence of lesions on the limbs and body were 93% (260/279) and 20% (57/288) respectively. The prevalence of limb and body lesions was significantly lower in outdoor-housed sows compared with indoor-housed sows. Indoor-housed sows had an increased risk of wounds (OR 6.8), calluses (OR 8.8) and capped hock (OR 3.8) on their limbs when housed on fully slatted floors compared with solid concrete floors. In addition, there was an increased risk of bursitis (OR 2.7), capped hock (OR 2.3) and shoulder lesions (OR 4.8) in sows that were unwilling to rise to their feet. There was a decreased risk of shoulder lesions (OR 0.3) and lesions elsewhere on the body (OR 0.2) in sows with more than 20 cm between their tail and the back of the crate compared with sows with less than 10 cm. Conclusion: The sample of outdoor housed sows in this study had the lowest prevalence of limb and body lesions. In lactating sows housed indoors there was a general trend for an increased risk of limb and body lesions in sows housed on slatted floors compared with those housed on solid concrete floors with bedding. Sows that were less responsive to human presence and sows that had the least space to move within their crates had an additional increased risk of lesions

A Cation–π Interaction between Extracellular TEA and an Aromatic Residue in Potassium Channels

Open-channel blockers such as tetraethylammonium (TEA) have a long history as probes of the permeation pathway of ion channels. High affinity blockade by extracellular TEA requires the presence of an aromatic amino acid at a position that sits at the external entrance of the permeation pathway (residue 449 in the eukaryotic voltage-gated potassium channel Shaker). We investigated whether a cation–{pi} interaction between TEA and such an aromatic residue contributes to TEA block using the in vivo nonsense suppression method to incorporate a series of increasingly fluorinated Phe side chains at position 449. Fluorination, which is known to decrease the cation–{pi} binding ability of an aromatic ring, progressively increased the inhibitory constant Ki for the TEA block of Shaker. A larger increase in Ki was observed when the benzene ring of Phe449 was substituted by nonaromatic cyclohexane. These results support a strong cation–{pi} component to the TEA block. The data provide an empirical basis for choosing between Shaker models that are based on two classes of reported crystal structures for the bacterial channel KcsA, showing residue Tyr82 in orientations either compatible or incompatible with a cation–{pi} mechanism. We propose that the aromatic residue at this position in Shaker is favorably oriented for a cation–{pi} interaction with the permeation pathway. This choice is supported by high level ab initio calculations of the predicted effects of Phe modifications on TEA binding energy

Testing the Impact of Higher Achievement's Year-Round Out-of-School-Time Program on Academic Outcomes

Presents findings from a multiyear evaluation of an intensive long-term OST program's effect on low-income middle school students' academic performance, attitudes, and behaviors. Outlines implications for financially strapped districts

A Cation-π Interaction Discriminates among Sodium Channels That Are Either Sensitive or Resistant to Tetrodotoxin Block

Voltage-gated sodium channels control the upstroke of the action potential in excitable cells of nerve and muscle tissue, making them ideal targets for exogenous toxins that aim to squelch electrical excitability. One such toxin, tetrodotoxin (TTX), blocks sodium channels with nanomolar affinity only when an aromatic Phe or Tyr residue is present at a specific location in the external vestibule of the ion-conducting pore. To test whether TTX is attracted to Tyr401 of NaV1.4 through a cation-{pi} interaction, this aromatic residue was replaced with fluorinated derivatives of Phe using in vivo nonsense suppression. Consistent with a cation-{pi} interaction, increased fluorination of Phe401, which reduces the negative electrostatic potential on the aromatic face, caused a monotonic increase in the inhibitory constant for block. Trifluorination of the aromatic ring decreased TTX affinity by ~50-fold, a reduction similar to that caused by replacement with the comparably hydrophobic residue Leu. Furthermore, we show that an energetically equivalent cation-{pi} interaction underlies both use-dependent and tonic block by TTX. Our results are supported by high level ab initio quantum mechanical calculations applied to a model of TTX binding to benzene. Our analysis suggests that the aromatic side chain faces the permeation pathway where it orients TTX optimally and interacts with permeant ions. These results are the first of their kind to show the incorporation of unnatural amino acids into a voltage-gated sodium channel and demonstrate that a cation-{pi} interaction is responsible for the obligate nature of an aromatic at this position in TTX-sensitive sodium channels

Summer Snapshot: Exploring the Impact of Higher Achievement's Year-Round Out-of-School-Time Program on Summer Learning

Assesses the impact of a multiyear, intensive, academically focused OST program for motivated but underserved middle school students on test scores, summer program participation, and summer learning loss. Examines contributing factors and implications

Use of an index to reflect the aggregate burden of long-term exposure to criteria air pollutants in the United States.

Air pollution control in the United States for five common pollutants--particulate matter, ground-level ozone, sulfur dioxide, nitrogen dioxide, and carbon monoxide--is based partly on the attainment of ambient air quality standards that represent a level of air pollution regarded as safe. Regulatory and health agencies often focus on whether standards for short periods are attained; the number of days that standards are exceeded is used to track progress. Efforts to explain air pollution to the public often incorporate an air quality index that represents daily concentrations of pollutants. While effects of short-term exposures have been emphasized, research shows that long-term exposures to lower concentrations of air pollutants can also result in adverse health effects. We developed an aggregate index that represents long-term exposure to these pollutants, using 1995 monitoring data for metropolitan areas obtained from the U.S. Environmental Protection Agency's Aerometric Information Retrieval System. We compared the ranking of metropolitan areas under the proposed aggregate index with the ranking of areas by the number of days that short-term standards were exceeded. The geographic areas with the highest burden of long-term exposures are not, in all cases, the same as those with the most days that exceeded a short-term standard. We believe that an aggregate index of long-term air pollution offers an informative addition to the principal approaches currently used to describe air pollution exposures; further work on an aggregate index representing long-term exposure to air pollutants is warranted

Land Management Decisions and Agricultural Productivity in the Hillsides of Honduras

Increasing land degradation and concomitant low agricultural productivity are important determinants of rural poverty in the hillside areas of Honduras. Using data at the levels of the farm household, parcel and plot, we develop an econometric modeling framework to analyze land management decisions and their impact on crop productivity. Our econometric model allows for endogenous household decisions regarding livelihood strategy choice, use of labor and external inputs, and participation in organizations. We found support for the inverse farm size-land productivity relationship which suggests that improved land access could increase total crop production. Land tenure has no impact on crop productivity, but adoption of soil conservation practices is higher on owner-operated than leased plots. Ownership of machinery and equipment and livestock ownership both positively influence crop productivity. Education positively affects perennial crop productivity. The gender of the household head has no significant effect on crop productivity, but does influence some land management and input use decisions. Even though household participation in training programs and organizations has only limited effects on crop productivity, agricultural extension plays a key role in promoting adoption of soil conservation practices. Location assets have limited impacts on crop productivity but do influence land management decisions. Road density and better market access have a positive effect on perennial crop productivity. Population density has limited direct impact on crop productivity, though it may have indirect effects by affecting farm size and livelihood strategies.agricultural productivity, hillsides, Honduras, land management, soil conservation, Land Economics/Use, Productivity Analysis,